In vitro biosynthesis and in vivo processing of the major microneme antigen of Sarcocystis muris cyst merozoites.

The cDNA clone pSM/1.6 encoding the 26.5-kDa precursor molecule of the 16/17-kDa microneme antigen of Sarcocystis muris cyst merozoites was expressed in a cell-free translation/translocation system to study translocation of the protein across membranes. The antigen was found to be translocated across heterologous endoplasmic reticulum membranes. Translocation was accompanied by cleavage of a signal peptide to create a 23-kDa polypeptide that was completely protected from digestion with proteinase K. Pulse-chase analysis of [35S]-methionine-labeled S. muris cyst merozoites demonstrated that the 16/17-kDa antigen derived from a 23-kDa precursor molecule and that its processing occurred at between a few minutes and 2 h after biosynthesis. This leads to the conclusion that the native microneme antigen is secreted from the parasite cell via the endoplasmic reticulum. Sorting into micronemes might occur during transition through a Golgi-like structure, involving cleavage of the hydrophilic propeptide to create the mature 16/17-kDa protein.